superarray product updates 2008

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SuperArray Product Updates 2008

Focus on Your Pathway

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Best Selling Product:

• Expression analysis tool for a focused panel of genes

• Pre-designed cataloged arrays for over 100 pathways

• Customized panels ranging from 4 to 384 genes

• Compatible with (almost) all real-time PCR instruments

• 96- or 384-well plate of SYBR Green-optimized qPCR assays

• 84 relevant, pathway- or disease-focused genes

• 5 housekeeping genes and three RNA quality controls

• Very simple two-hour protocol that generates:

• Microarray-like multi-gene content & results

• qPCR sensitivity and reproducibility

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RT2Profiler™ PCR Array Layout

G01 G02 G3 G04 G05 G06 G07 G08 G09 G10 G11 G12

G13 G14 G15 G16 G17 G18 G19 G20 G21 G22 G23 G24

G25 G26 G27 G28 G29 G30 G31 G32 G33 G34 G35 G36

G37 G38 G39 G40 G41 G42 G43 G44 G45 G46 G47 G48

G49 G50 G51 G52 G53 G54 G55 G56 G57 G58 G59 G60

G61 G62 G63 G64 G65 G66 G67 G68 G69 G70 G71 G72

G73 G74 G75 G76 G77 G78 G79 G80 G81 G82 G83 G84

HK1 HK2 HK3 HK4 HK5 GDC RTC RTC RTC PPC PPC PPC

• Wells A1-G12 (G01-G84): 84 Pathway-Focused Genes

• Wells H1-H5 (HK1-HK5): 5 Housekeeping Genes

• Wells H6 (GDC): Genomic DNA Control

• Well H7-H9 (RTC): Reverse Transcription Control

• Well H10-H12 (PPC): Positive PCR Control

1 2 3 4 5 6 7 8 9 10 11 12

A B

C D

E F

G H

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Market Acceptance of PCR Array

• Launched May 2005

• V2.0 November 2006, V3.0 September 2007

• Better Controls, Up-to-Date Sequence Info

• Over 2000 Customers Worldwide

• Current Publication List >200, 30+ for PCR Array

• Most added in last 3 months of 2007

• Expected to grow very rapidly

• See how researchers like you have used SuperArray technologies in their research [ Go ]

• Conferences / Tradeshows

• More & More Posters with Data Using PCR Arrays

• More Customers stopping by & pleased with PCR Array

http://www.superarray.com/support_publication.php

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Customer Success Story

" The use of the RT2 Profiler Human Th1-Th2-Th3 PCR array has provided important insight into the effects of cocaine and the results of the mRNA profiling have correlated well with additional work on cocaine-mediated changes in cytokine secretion and T cell function"

Baldwin GC, Professor and Director, UCLA

Presented at the 2008 Meeting of The College on Problems of Drug Dependence.

http://www.uclaaidsinstitute.org/index.html

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Baylor College of Medicine

Baylor College of Medicine- Feigin Center

Stanford University

University of Washington

Thomas Jefferson Kimmel Cancer Center

Trudeau Institute

University of Pennsylvania

UCSF

UCLA

The University of Texas Health Science Center-

Houston

Technology Leaders’ Endorsement

PCR Array Core Facilities Program in US

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New PCR Array Pathways

• Brand New Pathways Coming Soon

• Aging and Senescence

• Asthma

• Autoimmunity & Self Tolerance

• Cell Membrane Receptors

• Cell Migration

• Cell Motility

• Circadian Pathways

• Cytoskeleton

• Cytotoxicity

• Gap Junction Proteins

• Genome Stability & DNA Repair

• Glucocorticoid Signaling

• G-Protein Coupled Receptors

• Hedgehog Signaling

• Immunomodulation

• Integrin Signaling

• Ion Channels

• Kinases

• Lipoprotein Signaling & Cholesterol Metabolism

• Membrane Trafficking

• Metal & Ion Transport

• miRNA Pathway

• Mitochondrial Function

• Neoplastic Tissue

• PKC & Phospholipase Signaling

• Prostate Cancer

• Proteases & Proteinases

• Protein Hydroxylases

• Protein Trafficking

• Proteoglycans

• RNA Interference Pathway

• Signal Transduction Scaffolding Proteins

• Tissue & Cell Regenerative Pathway

• Ubiquitin Degradation

• Vascular Development Pathway

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Useful Resources for Gene Expression Researchers-Forward to your friends!

http://www.superarray.com/seminarlist.php

http://www.superarray.com/support_faq.php

Real-time PCR

RNA interference

-provide you insights on

these topics based on

our expertise

PCR Array

Analysis Portal

-useful web-based

software for real-time

PCR data analysis


http://www.superarray.com/pcr/arrayanalysis.php

http://www.superarray.com/support_posters.php

PostersPublications

http://www.superarray.com/pathwaycentral.php

Pathway Central NEW!

- find pathway reviews

and presentation-ready

pathway maps

-overview of SuperArray core technologies







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New product 2008:

1) RT2 miRNA PCR Array

2) Cignal Reporter Assays

3) ChIP-qPCR Assays

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1) RT2 miRNA qPCR Array & Assay System

Sensitivity:

As little as 0.5 µg total RNA needed

Multi-Sequence Flexibility:

Analyze up to 376 sequences simultaneously

Simplicity:

As easy as a real-time PCR experiment

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How the miRNA PCR Array Works

Control & Experimental Cells or Tissue

1. Isolate miRNA (MA-01)

2. Reverse Transcription: RT2 miRNA First Strand Kit (MA-03)

3. Setting Up the PCR Array

Instrument-Specific RT2 qPCR SYBR Green Master Mix & cDNA Cocktail

Aliquot across PCR Array

4. Real-Time PCR

(40 cycles, 100-120 min)

5. Data Analysis: Determine Fold Changes (DD Ct Method)

Excel Template & Web Portal NOW Available

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Small RNA Enrichment

TRIzol Aqueous Phase

Adjust EtOH Conc.

Small RNA Elutes

Large Nucleic Acid Binds

Readjust Eluate

Adjust EtOH Conc.Small RNA binds

Wash

Elute Small RNA

Appendix Protocol

Purifies Large RNA

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RT2 miRNA First Strand Kit (MA-03)

M1: miRNA RT Primer & ERC Mix

M2: 5X miRNA RT Buffer

M3: miRNA RT Enzyme Mix

RNase-free H2O

TTTTTTTT

miRNA

AAAAAAAATTTTTTTT

miRNA RT Primer

Reverse Transcriptase

Poly(A) Polymerase

Real-Time PCR

TTTTTTTT

Gene-Specific Primer

Universal Primer

Universal Reverse Transcription Method:

Same cDNA preparation can be used for ANY gene

Single-Tube 30-min Reaction

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miRNA PCR Array Performance

miRNA ID Sequence

miR-10a UACCCUGUAGAUCCGAAUUUGUG

miR-10b UACCCUGUAGAACCGAAUUUGUG

miRNA-99a AACCCGUAGAUCCGAUCUUGUG

miRNA-100 AACCCGUAGAUCCGAACUUGUG

miR-196a UAGGUAGUUUCAUGUUGUUGGG

miR-196b UAGGUAGUUUCCUGUUGUUGGG

Relative PCR cross-reactivity is calculated from the difference in Ct values

between using target- or isoform-specific primers to detect 105 copies of

specific miRNA target cDNA template.

Specificity / Discrimination

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miRNA PCR Array Performance

The proprietary primer design of the RT2 miRNA PCR Arrays and Assays

distinguishes miRNA family members with single nucleotide mismatches.

Specificity / Discrimination

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Cancer Application Data

Small RNA isolated from human colon tumor and matched adjacent normal

tissue (Biochain) was characterized with on the Human Cancer RT2 miRNA

PCR Array. Many cancer-specifc miRNA are up-regulated in this model.

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2) Cell-based Cignal Reporter Assays

• Product Line Breadth

• 18 Pathway-Focused Reporter Assays

• 4 Application-Specific 10-Pathway Suites

• Performance

Exceptional Sensitivity, Specificity, Signal-to-Noise Ratio

• Convenience

Ready-to-use, preformatted transient assays

• Dual Modalities

• Dual-luciferase quantitative endpoint assays

• GFP dynamic live cell assays with single cell resolution

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Cignal Applications

• Functional Genomics – What’s the phenotype of my gene?

Does my gene of interest regulate signaling pathway “X”?

• Functional Proteomics – What’s the phenotype of my protein/peptide?

Does my protein, peptide, or growth factor of interest regulate signaling

pathway “X”?

• Drug Screening – What is the mechanism of action of my small molecule

drug candidates?

Do my drug candidates of interest regulate signaling pathway “X”?

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REPORTER DESIGN: Dual-Luciferase Assays

Reporter Construct Firefly luciferaseTATA

boxTRE

Renilla luciferaseTATA

boxCMVNormalization Construct

Pathway-targeted transcriptional

regulatory elements (18)

Constitutive transcriptional

regulatory element (1)

The Transcriptional Regulatory Elements (TREs)

are what establish the

pathway specificity of each reporter

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How Does the Cignal Reporter Assay Work?

We have

experimentally

optimized the TREs

to maximize the

signal-to-noise

ratio

3 components:

1) p/w transcription factor-

responsive firefly

luciferase reporter

2) Non-inducible firefly

lucifirase -negative

control

3) Mixture of constitutively

expressing GFP and

firefly lucifierase-

positive control

Constitutively

expressing Renilla

luciferase reporter as

normalizing. construct

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Cignal Reporter Assays

Pathway

C/EBP

cAMP/PKA

Cell Cycle

DNA Damage

Estrogen Receptor

Glucocorticoid Receptor

Hypoxia

Type I Interferons

Interferon Gamma

MAPK/ERK

MAPK/JNK

c-Myc

NFkB

Notch

PKC/Ca++

Retinoic Acid Receptor

TGFβ

Wnt

Transcription Factor

C/EBP

CREB

E2F/DP1

p53

Estrogen Receptor


HIF

STAT1/STAT2

STAT1/STAT1

Elk-1/SRF

AP1

Myc/Max

NFkB

RBP-Jk

NFAT

Retinoic Acid Receptor

SMAD2/SMAD3/SMAD4

TCF/LEF

Regulatory Element

C/EBP binding site

CRE

E2F binding site

p53 response element

ERE

GRE

HIF response element

ISRE

GAS

SRE

AP1 binding element

E-box binding element

NFkB binding element

RBP-Jk response element

NFAT response element

RARE

SMAD response element

TCF/LEF response element

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Cignal Cancer Suite

Pathway

Cell Cycle

DNA Damage

Hypoxia

MAPK/ERK

MAPK/JNK

c-Myc

NFkB

Notch

TGFβ

Wnt


E2F/DP1

p53

HIF

Elk-1/SRF

AP1

Myc/Max

NFkB

RBP-Jk

SMAD2/SMAD4

TCF/LEF

Regulatory Element

E2F binding site



SRE

AP1 binding element






Cignal Cancer Reporter Assay 10-Pathway Suite (luc) CCA-001L

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Cignal Immune Response Suite

Pathway

C/EBP

cAMP/PKA


Type I Interferons

Interferon Gamma

MAPK/ERK

MAPK/JNK

NFkB

PKC/Ca++

TGFβ


C/EBP

CREB


STAT1/STAT2

STAT1/STAT1

Elk-1/SRF

AP1

NFkB

NFAT

SMAD2/SMAD4

Regulatory Element

C/EBP binding site

CRE

GRE

ISRE

GAS

SRE

AP1 binding element




Cignal Immune Response Reporter Assay 10-Pathway Suite (luc) CCA-002L

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Cignal Growth & Differentiation Suite

Pathway

C/EBP

cAMP/PKA

Cell Cycle

MAPK/ERK

MAPK/JNK

c-Myc

NFκB

Notch

TGFβ

Wnt


C/EBP

CREB

E2F/DP1

Elk-1/SRF

AP1

Myc/Max

NFκB

RBP-Jk

SMAD2/SMAD4

TCF/LEF

Regulatory Element

C/EBP binding site

CRE

E2F binding site

SRE

AP1 binding element


NFκB binding element




Cignal Growth & Diff. Reporter Assay 10-Pathway Suite (luc) CCA-003L

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Cignal Stress & Toxicity Suite

Pathway

Cell Cycle

DNA Damage


Hypoxia

MAPK/ERK

MAPK/JNK

c-Myc

NFkB

PKC/Ca++

TGFβ


E2F/DP1

p53


HIF

Elk-1/SRF

AP1

Myc/Max

NFkB

NFAT

SMAD2/SMAD4

Regulatory Element

E2F binding site


GRE


SRE

AP1 binding element





Cignal Stress & Toxicity Reporter Assay 10-Pathway Suite (luc) CCA-004L

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3) ChIP-qPCR Array and Assay System

“A ChIP Off The Old (Real-Time PCR) Block”

Speed & Ease:

ChIP Antibody & qPCR Assay already optimized and ready to use

Comprehensive:

Portfolio will eventually represent the 20 most popular ChIP targets – including

transcription factors & histone modifications

High Performance:

Qualified ChIP Antibody & qPCR Assay provide positive specific signals in a well-

defined cell-based experiment when used with our ChIP Prep Kit

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HOW IT WORKS: Tiling Scheme

• Many transcription factor binding sites (TF BS) per promoter

• Some known, some yet to be discovered

• Optimal qPCR primer design with stringent criteria needs room

to find best amplicon sequence

• Localizing amplicon near TF BS can be a hindrance

• On average, amplicons located within 500 bp from TFBS

• Most fragmented chromatin about that size anyway

qPCR Assay & Amplicon Amplicon Coverage Range (Tile)

Proximal Promoter

TSS (+1)mRNA

gDNA

Structural Gene

Distal Promoter

TF BS

1 kb 1 kb1 kb 1 kb1 kb

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ChIP-qPCR Search Portal

Gene Symbol:

Transcription Factor:

AND / OR

ChIP-on-chip / aCGH Validators OR CNV-qPCR Users

Chromosome RefSeq:

Position:

New to ChIP? Find your Transcription Factor or Gene.

Currently using ChIP? Find a new assay for your queried target.

http://www.superarray.com/chipqpcr.php

SUBMIT

SUBMIT

Any TF ▼

http://www.superarray.com/chipqpcr.php

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Application Example

• Treat four human cancer cell lines (A549, HepG2, MCF7, PC3) with anti-cancer agent 5-flourouracil (5-FU)

• Grow & Treat Cells with Biological Replicates

• Monitor p53 signaling pathway gene expression

• Human p53 Signaling RT2 Profiler™ PCR Array

• Verify increases in gene expression with p53 ChIP-qPCR

• Formaldehyde Cross-Link; Lyse Cells & Prepare Chromatin

• Fragment DNA by Sonication to 800-1,000-bp Average Size

• ChIP with Human p53 & RNA Pol II Target-Specific Kits

Plus Non-Immune Serum negative control

• Measure p53 & RNA Pol II enrichment at gene promoters

Plus Negative & Positive Control qPCR Assays

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Application Example Results

31

Useful Resources for Gene Expression Researchers-Forward to your friends!



Real-time PCR

RNA interference

-provide you insights on

these topics based on

our expertise

PCR Array

Analysis Portal

-useful web-based

software for real-time

PCR data analysis




PostersPublications


Pathway Central NEW!

- find pathway reviews

and presentation-ready

pathway maps

-overview of SuperArray core technologies







superarray product updates 2008

Documents